Phyto-gene therapy using antisense oligonucleotides to control cereal fungal disease by silencing virulence factors and their regulators

With increasing concerns regarding food security, alternative solutions are required for disease control in crops, including those caused by fungal pathogens. Antisense single stranded short oligodeoxynucleotides (ASO) based gene therapy is widely used in medicine but is still emerging in plant sciences. The ASO gene silencing approach using phosphorothioate modified oligodeoxynucleotides (asPTOs) delivered to excised barley leaves was first devised as a tool for in planta transient host induced gene silencing (HIGS) to query the virulence role of genes from the biotrophic fungal pathogen, Blumeria graminis f.sp. hordei (Bgh), the causal agent of barley powdery mildew.
Following this, our project aimed at exploiting the HIGS approach for discovering new key players for virulence of Bgh and some of the major wheat pathogens, B. graminis f.sp. tritici (Bgt) and Fusarium graminearum, the causal agent of Fusarium head blight. The ASO gene silencing approach was also evaluated for its suitability to protect wheat against fungi by targeting host susceptibility genes.
AsPTOs to silence vital Bgh genes (actin, GAPDH, 2-Glycosyl transferase) successfully reduced powdery mildew infection in several barley cultivars. Similarly, silencing the metallo-protease-like effector BEC1019 impacted on Bgh and Bgt virulence in barley and wheat respectively. Following promoter sequence analysis of Bgh effectors expressed in haustoria, the HIGS approach allowed to confirm the implication of ZAP1 and PacC transcription factors in regulating BEC1019 and BEC1011 effector expression, while affecting Bgh virulence.
To adapt ASO based gene silencing for disease control, in planta gene silencing of F. graminearum known virulence genes was attempted but with no convincing impact. However, asPTOs to silence BEC1011 were delivered into whole barley seedlings by root uptake resulting in reduced powdery mildew infection. This suggests that asPTOs based HIGS could be further investigated as a strategy to control fungal diseases in crops.